Abstract
Non-melanoma skin cancer (NMSC) is the most common form of cancer. Ultraviolet-B (UVB) radiation has been shown to be a complete carcinogen in the development of NMSC. The mammalian target of rapamycin complex 1 (mTORC1) is upregulated by UVB. Ornithine decarboxylase (ODC), the first enzyme of the polyamine biosynthetic pathway, is also upregulated in response to UVB. However, the interplay between these two pathways after UVB exposure remains unclear. The studies described here compare mRNA stability between normal human keratinocytes (HaCaT cells) and HaCaT cells with low levels of raptor to investigate whether the induction of ODC by UVB is dependent on mTORC1. We show that the knockdown of mTORC1 activity led to decreased levels of ODC protein both before and after exposure to 20 mJ/cm2 UVB. ODC mRNA was less stable in cells with decreased mTORC1 activity. Polysome profiles revealed that the initiation of ODC mRNA translation did not change in UVB-treated cells. We have shown that the ODC transcript is stabilized by the RNA-binding protein human antigen R (HuR). To expand these studies, we investigated whether HuR functions to regulate ODC mRNA stability in human keratinocytes exposed to UVB. We show an increased cytoplasmic localization of HuR after UVB exposure in wild-type cells. The ablation of HuR via CRISPR/Cas9 did not alter the stability of the ODC message, suggesting the involvement of other trans-acting factors. These data suggest that in human keratinocytes, ODC mRNA stability is regulated, in part, by an mTORC1-dependent mechanism after UVB exposure.
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Acknowledgements
The authors would like to thank Dr. Scot Kimball (Penn State College of Medicine) for his help with the polysome profiles. Work in the authors’ laboratory was funded by grants from the National Institutes of Health (ES19242 to LMS, ES26471 to RPF) and funds from Penn State Berks to SLN.
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Stump, C.L., Feehan, R.P., Jordan, T. et al. Knocking down raptor in human keratinocytes affects ornithine decarboxylase in a post-transcriptional Manner following ultraviolet B exposure. Amino Acids 52, 141–149 (2020). https://doi.org/10.1007/s00726-019-02732-3
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DOI: https://doi.org/10.1007/s00726-019-02732-3